The present invention relates to a diagnostic composition for detecting an affective disorder, to methods utilizing the same and to kits containing the same. More particularly the present invention relates to compositions, methods and kits for diagnosis and monitoring of treatment of affective disorders.
Mental disorders with their very high prevalence and incidence represent a significant share of all causes of disability worldwide. In 1990, the World Health Organization (WHO) have calculated that of the ten leading causes of disability worldwide four are mental disorders and the number one cause is unipolar major depression (Table 1).
TABLE 1Leading Causes of Disability Worldwide, World Health Organization1990Percent ofTotal (millions)total1. Unipolar major depression50.810.72. Iron-deficiency anemia22.04.73. Falls22.04.64. Alcohol use15.83.35. Chronic pulmunary disease14.73.16. Bipolar disorder14.13.07. Congenital anomalies13.52.98. Osteoarthritis13.32.89. Schizophrenia12.12.610. Obsessive-compulsive disorder10.22.2All causes472.7
Major depression ranked fourth, behind lower respiratory infections, diarrheal diseases, and perinatal conditions, in terms of disease burden measured in disability-adjusted life years. The National Co-morbidity Study, a large scale epidemiological survey of mental illness prevalence, indicated that the life time prevalence of major depression is 17%. The survey also indicated that in any given year 10% of the individuals, aged 18-54 years, suffer from a depressive disorder.
Most of the present modalities of pharmacological treatments in mental disorders already exist for about 50 years. Lithium, the drug of choice for the treatment of bipolar mood disorder was discovered to possess anti-manic therapeutic potential by John Cade in 1949. The first anti-psychotic medication, chlorpromazine was discovered by Delay & Deniker in 1952. Similarly the first tricyclic antidepressant, imipramine, as well as the first benzodiazepine anti-anxiety drug, chlordiazepoxide, were discovered in 1957 (Table 2).
TABLE 2Modalities of PsychopharmacologicalTreatments Discovered in the 1950sTreatmentDiscoverer ofModalityPrototypicalMedicationYearMood stabilizerLithiumJ. F. J. Cade1949Anti-psychoticChlorpromazineJ. Delay & P. Deniker1952Anti-depressant:TricyclicImipramineR. Kuhn1958MAO-inhibitorIproniazidN. S. Kline1958Anti-anxietyChlordiazepoxideRandell1957
From that bifurcating time point in the history of psychopharmacology only few new treatment modalities i.e., anti-obsessive-compulsive medications, have appeared, and only few improvements in clinical efficacy of the existing treatment modalities have came upon. Based on pathophysiological hypotheses, new drugs (e.g. serotonin-selective re-uptake inhibitors as new antidepressants, dopamine-serotonin antagonists as new anti-psychotics) have been developed. The most notable improvements exist in the development of medications with fewer side effects.
The existence of biological-pharmacological treatments in mental disorders has appropriated psychiatry as a medical discipline. The medical model has become the dominant mode of psychiatry world-wide. In this model psychiatrists are seen as physicians who specialize in the treatment of mental disorders. Diagnosis is used to predict the future course of a disorder and its likely response to treatment, and it is the foundation for all subsequent therapeutic decisions. In contrast, the adaptation model used by psychoanalytic psychiatry for example sees psychiatrists as specialists in behavior and adaptation whose expertise can benefit people whether or not they have a diagnosable mental disorder.
There is a significant gap between advances in medication for mental disorders and the present static situation in diagnosis and monitoring treatments of these disorders. As with all areas of medicine, treatment decisions are guided by diagnosis. Unlike most disciplines of physical medicine, psychiatry has no external validating criteria. The present categorical classification of mental disorders and, accordingly, differential diagnosis in Psychiatry, divides mental disorders into types based on clusters of clinical signs, symptoms and their time course. The current Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) of the American Psychiatric Association and International Statistical Classification of Diseases and Related Health Problems classification of mental disorders (ICD-10, Chapter V), developed by the World Health Organization, both use an empirical, non-theoretical orientation and operational criteria, which are predominantly phenomenological. These classification systems use explicit diagnostic criteria, and a descriptive approach that attempts to be neutral with respect to theories of etiology (e.g. biological psychiatry, psychoanalysis). DSM and ICD categorical classification divides mental disorders into types based on clusters of clinical signs, symptoms and their time course. Table 3 exemplifies DSM-IV criteria for major depressive episode.
TABLE 3DSM-IV Criteria for Major Depressive Episode1. Five (or more) of the following symptoms have been present during thesame 2-week period and present a change from previous functioning; atleast one of the symptoms is either (1) depressed mood or (2) loss ofinterest or pleasure.(1)Depressed mood most of the day, nearly every day, as indicated byeither subjective report (e.g., feels sad or empty) or observationmade by others (e.g., appears tearful).(2)Markedly diminished interest or pleasure in all, or almostall, activities most of the day, nearly every day.(3)Significant weight loss when not dieting or weight gain (e.g.,a change of more than 5% of body weight in a month), or decreaseor increase in appetite nearly every day.(4)Insomnia or hypersomnia nearly every day.(5)Psychomotor agitation or retardation nearly every day.(6)Fatigue or loss of energy nearly every day.(7)Feelings of worthlessness or excessive or inappropriate guilt nearlyevery day.(8)Diminished ability to think or concentrate, or indecisiveness, nearlyevery day.(9)Recurrent thoughts of death, recurrent suicidal ideation without aspecific plan, or a suicidal attempt or a specific plan for committingsuicide.1. The symptoms do not meet criteria for a Mixed Episode (Manic andMajor Depressive Episode).2. The symptoms cause clinically significant distress or impairment insocial, occupational, or other important areas of functioning.3. The symptoms are not due to the direct physiological effects of asubstance or a general medical condition.4. The symptoms are not better accounted for by Bereavement.
In contrast to the exciting advances made in the psychopharmacology of mental disorders and in spite of a considerable research effort directed at attempting to decipher the biological underpinnings of psychiatric disorders, no biochemical cross-diagnostic tests capable of supporting psychiatric differential diagnosis and treatment monitoring are available. Although the structured algorithms described in the DSM and ICD diagnostic systems have been extensively tested and revised to improve their reliability and validity, these diagnostic criteria rely upon subjective reports by the patients and subjective elements in their evaluation by psychiatric specialists. There are presently no reliable, sensitive and specific objective biological diagnostic markers that can serve as ‘gold standards’.
The importance of a biochemical assay for major depression lays in the ability, both by the general practitioner, who treats about half of these patients, and by mental health professionals, to decide on the application and follow up of biological antidepressant treatments. While it is recognized that significant morbidity and mortality is associated with depression, failure to appropriately diagnose individuals remains the primary obstacle to the effective management of this disorder. The results of the NIMH Epidemiologic Catchment Area program, which assessed mental disorder prevalence, incidence and service use in about 20,000 community and institutional residents have shown that there is limited treatment of individuals in the community diagnosed with psychiatric disorders. Concerning mood disorders, for example, only 11% of individuals with major depression had received an antidepressant medication. When individuals with major mental disorders seek treatment, they are usually underdiagnosed, or if correct diagnosis is made, they receive inadequate psychopharmacological intervention. More than 50% of the population who seek treatment for mental disorders usually turn to general practitioners and other physicians other than psychiatrists. Less than 50% of those who turn to general practitioners for depressive illness are diagnosed properly. Less than 50% of those who are properly diagnosed by the general practitioner receive the proper medicative treatment. Less than 50% of those who do receive proper pharmacological treatment by the general practitioner, receive the proper dosage.
The Medical Outcomes Study, a four-year longitudinal study of more than 20,000 patients with chronic conditions in different practice settings in the United States, found that depressed patients who consulted a family physician in a fee-for-service setting had roughly a 55% chance of having their condition diagnosed correctly. The detection rate for patients consulting primary care physicians in managed care organizations was even lower: approximately 40%.
The serendipitous discovery of antidepressant medications more than 50 years ago provided the first evidence of an inherent biochemical abnormality underlying the disorder. The ‘pharmacocentric’ approach to the construction of biological hypotheses for pathogenesis of mental disorders is trying to induce from knowledge and hypotheses concerning the biochemical mechanism of action of neuropsychiatric medications to pathogenic and pathophysiologic hypotheses. Research into the mechanism of action of the first tricyclic antidepressant imipramine and the first monoamine oxidase inhibitor-type of antidepressant iproniazide, has shown that increasing synaptic concentrations of monoamines (re-uptake inhibition of serotonin and/or norepinephrine to presynaptic vesicles;
monoamine oxidase inhibition) can improve the symptoms of depression. Thus a role for monoaminergic mechanisms in the pathogenesis of mood disorders has been formulated in the catecholamine, the serotonin, and the monoaminergic-cholinergic balance hypotheses of, mood disorders. The monoaminergic hypothesis did not provide an adequate explanation to the lag period of 10-20 days in the therapeutic actions of antidepressants. Thus, in the mid-1970s, theories on the mode of action of antidepressants shifted the emphasis from acute pre-synaptic events to delayed post-synaptic adaptations at the level of receptors.
Antidepressants are well known for their potential to modulate the density of functional neurotransmitter receptors such as beta adrenergic receptors in the brain, as well as in cultured cells. Beta-adrenergic down-regulation is accompanied by decreased receptor-stimulated cAMP formation. The mechanisms for this reduction in receptor numbers are not completely understood. Importantly, the onset of down-regulation and clinical effectiveness requires 10-20 days of antidepressant treatment. It has been proposed that the reduction in the number of functional beta-adrenergic and other receptors could be a regulatory response to the enhanced presence of the neurotransmitter in the synaptic cleft. The neurotransmitter's elevated concentration is induced by acute inhibition of its reuptake or of monoamine oxidase activity by antidepressants. Some clinically effective antidepressants, however, neither influence norepinephrine or serotonin reuptake nor inhibit monoamine oxidase activity but still cause receptor down-regulation. Furthermore, this model fails to explain the observed time lag between the rapid drug-induced increase in intra-synaptic neurotransmitter concentrations and the delayed receptor down-regulation.
Decreased monoamine receptors densities following antidepressant treatment can also be seen in cell culture systems lacking a presynaptic input. Thus, monoamine receptor down-regulation may directly result from post synaptic actions of the antidepressants. Very recent studies suggest that antidepressants might induce switch of beta-adrenergic receptor trafficking as a mechanism for their action.
It has recently been found that there are signal transduction components involved in receptor desensitization.
Almost all G protein coupled receptors are tightly regulated by a common desensitising mechanism. The process of agonist-specific, homologous desensitization of receptors is characterized by an increase in the refractoriness of a receptor to signal in response to repeated or sustained exposure to its agonist, limiting both the magnitude and the temporal extend of the receptor signal, thus protecting cells from over-stimulation. The waning of G-protein-coupled-receptor (GPCR) signaling in the continued presence of agonist is accomplished by a coordinated series of events that are typically considered as three distinct processes:                (i) receptor desensitization—mediated by uncoupling the activated receptor from its specific G protein and achieved within a few minutes following agonist exposure;        (ii) receptor sequestration or endocytosis—mediated by receptor internalization from the plasma membrane to early endosomes, a process that occurs more slowly than desensitization, happening over a period of several minutes to hours after agonist exposure;        (iii) down-regulation—receptor degradation, a process that leads to a reduction in the total number of receptors.        
The mechanism underlying the phenomenon of desensitization involves the activities of two families of proteins: G protein-coupled receptor kinases (GRKs), and arresting. GRKs comprise a highly regulated cytosolic, multigene family of serine-threonine kinases, capable of specifically phosphorylating the agonist-occupied form of GPCRs. Receptor phosphorylation by GRKs has been ultimately identified as the initial and critical step in the uncoupling of receptor from G protein leading to the attenuation or desensitization of GPCR signaling. GRKs are translocated to the plasma membrane for their appropriate interaction with receptor domains. It is known that free G protein βγ subunits bind to the C-terminal domain of GRK and facilitate the translocation process. Today seven GRKs have been identified, two of them specific to the visual system (GRK1 & 7), one selectively present in sperm cells (GRK4), and four ubiquitously distributed.
Activated heptahelical receptors are phosphorylated by a family of G protein-coupled receptor kinases (GRKs). Following phosphorylation, the receptors bind to another family of proteins called arresting. Arrestin is the name given to a class of soluble proteins that are found to stop or ‘arrest’ intracellular signaling triggered by G protein coupled transmembrane receptors. The regions of the receptors that arrestins bind to, are also primary determinants for G protein interaction. Arrestin binding to receptors thus results in desensitization of G protein-mediated signaling by preventing interaction of receptors with G proteins. To date, four members of the arrestin gene family have been cloned. Two arrestins, visual arrestin and cone arrestin, are expressed almost exclusively in the retina, where they regulate photoreceptor function, while β-arrestin1 and β-arrestin2, are ubiquitously expressed proteins. Although most of the research regarding the desensitization process has been carried out using the beta-2-adrenergic receptor as a model, it is now clear that this process regulates the function of many GPCRs, including α- and β-adrenergic, muscarinic cholinergic, serotonergic, dopamine, angiotensin, endothelin, etc.
An emerging view, however is that the binding of arrestins to heptahelical receptors also initiates a new set of signaling pathways in addition to blocking those mediated by G protein activation. The association of arrestins with heptahelical receptors does not simply uncouple receptors from G protein pathways, but rather induces a switch in receptor signaling from classical second messenger-generating G protein-mediated pathways to other pathways such as those involving tyrosine kinases of the Src family, and leading to activation of MAP kinase. Moreover, arrestins have also been found to interact with a number of cellular proteins involved in endocytosis such as clathrin. Thus arrestins may well represent multifunctional adaptor proteins that mediate a number of aspects of heptahelical receptor signaling. GRKs may also be signaling intermediates for heptahelical receptors rather than just proteins involved in receptor desensitization. Recently, it was found that GRK2 can associate with actin and tubulin and can phosphorylate tubulin. Thus the recruitment of GRKs to activated heptahelical receptors may lead to cytoskeletal regulation or to modulation of other intracellular processes.
A differential pattern of receptor-coupled G protein measures was detected in mononuclear leukocytes of patients with the major mental disorders of mania, depression, schizophrenia, and panic disorder. G protein measures were suggested as a possible differential diagnostic test for these disorders [Table 4].
TABLE 4Differential G Protein Measures in Leukocytes of Patients with Mental DisordersDisorderG protein functionG protein levelG protein alteredmeasureAgonist - enhancedImmunoreactivityNormalization byGuanine nucleotidespecific treatmentbindingManiaIncreased β-GsIncreased GαsLithiumIncreased M-GiIncreased GαlLithiumDepressionDecreased β-GsDecreased GαsAntidepressants, ECTDecreased M-GiDecreased GαiAntidepressants, ECTWinter DepressionDecreased GαsLight therapyIn SADDecreased GαlLight therapySchizophreniaNormal-like β-GsNormal-like GαsAntipsychoticsNormal-like M-GiNormal-like GαlAntipsychoticsIncreased D-GsAntipsychoticsPanicIncreased β-GsNormal-like GαsDecreased M-GiNormal-like Gαiβ-Gs: β-adrenergic receptor coupled Gs protein function;M-Gi: Muscarinic receptor coupled Gi protein function;D-Gs: Dopamine receptor coupled Gs protein function.[for review see Avissar & Schreiber, 2002; Schreiber & Avissar, 2003]. G protein measures were also applied as an objective measure of depression in patients with irritable bowel syndrome.
The ability to detect state-dependent, disorder-specific alterations in G protein measures in patients with major depressive disorder, mania, panic disorder and schizophrenia, supported the use of these measures to biochemically monitor normalization of altered G protein measures during specific treatments [Table 4]. Indeed the elevated function of Gs and Gi proteins detected in patients during a manic episode was normalized by lithium treatment. Reduced Gs and Gi proteins level and function in patients during a depressive episode were corrected by antidepressants and ECT. The dynamics of normalization by electroconvulsive treatment of the biochemical measures of Gs and Gi function and of Gαs and Gαi levels was found not to follow and thus reflect the clinical improvement of the depressed patients, but rather to precede clinical improvement. The biochemical findings preceded clinical improvement by at least 1½ week. In SAD patients the extent of normalization of G protein measures by light therapy segregated well between responders and non-responders to light therapy.
Use of the state dependency of G protein measures in mood disordered patients was suggested as a mean to monitor and predict response to antidepressant treatments. It is very difficult to monitor the extent of clinical response especially in the early period of the first and second week after initiation of antidepressant treatment. Clinical response to antidepressant treatments is due both to the specific biochemical antidepressant effects of the medication, as well as to placebo effects. Since the placebo effect is usually more pronounced during the early period of treatment initiation, it is very difficult to assess in these early days the specific antidepressant effects of antidepressant treatments. G protein measures in peripheral blood elements of patients with mood disorders, as a state dependent characteristic, may afford biochemical monitoring and prediction of clinical response.
Dopamine-induced Gs hyperfunction characteristic of untreated patients with schizophrenia was corrected by treatment with either the classical anti-psychotic haloperidol and the newer anti-psychotic clozapine [Table 4]. Haloperidol caused a significant decrease in Gs level and function to below normal levels, characteristic of patients with Parkinson's disease. The extend of reduction in Gs measures was found to be correlated with the intensity of the Parkinsonian side effects, enabling the biochemical monitoring not only of anti-psychotic effects, but also of the extend of parkinsonian side effects.
Thus, according to the present invention there is now provided a diagnostic composition for detecting an affective disorder comprising an antibody selected from the group consisting of an antibody to a beta-arrestin, an antibody to a G-protein coupled receptor kinase and combinations. thereof as the active detecting agent therein.
In preferred embodiments of the present invention there is provided a diagnostic composition for detecting depression comprising an antibody selected from the group consisting of an antibody to a beta-arrestin, an antibody to a G-protein coupled receptor kinase and combinations thereof as the active detecting agent therein.
The invention also provides a composition for monitoring the effectiveness of an antidepressant, said composition comprising an antibody selected from the group consisting of an antibody to a beta-arrestin, an antibody to a G-protein coupled receptor kinase and combinations thereof as the active monitoring agent therein.
In another aspect of the present invention there is provided a method for diagnosing an affective disorder, or gauging the effect of a treatment upon a patient suffering from an affective disorder comprising determining the level of a beta-arrestin present in a patient and diagnosing the affective disorder or gauging the effect of the treatment upon the patient based on said determination.